Nitrogen is an essential plant nutrient which is required for the biosynthesis of nucleotides for DNA and RNA and amino acids for proteins. Although nitrogen composes nearly 78% of Earth's atmosphere, the majority is diatomic (N2), which is not a useful form for plants. Although useful nitrogen compounds (e.g., ammonia) are widely produced through industrial chemical means as fertilizers, it is estimated that more than 4/5 of all useful nitrogen compounds produced on an annual basis are produced naturally, the vast majority of which is produced by diazotrophic microorganisms.
Diazotrophs are microorganisms that have the ability to convert diatomic nitrogen into nitrogen-containing compounds which can be used by plants. This process is known as biological nitrogen fixation, or simply nitrogen fixation. Biological nitrogen fixation can be modeled by the following chemical equation:(N2+8H++8e−+16ATP=2NH3+H2+16ADP+16Pi)Thus, during nitrogen fixation, two moles of ammonia are produced from one mole of nitrogen at the cost of sixteen moles of adenosine triphosphate (ATP).
Diazotrophs can be divided into two categories: symbiotic and free-living. Symbiotic diazotrophs form associations with plants or animals. For example, Bradyrhizobium japonicum is a gram negative, rod-shaped, nitrogen-fixing bacterium which develops a symbiosis with soybeans. B. japonicum belongs to the family, Rhizobiaceae which includes other nitrogen-fixing bacteria that develop symbioses with legumes. In a symbiotic relationship, the bacteria produce fixed nitrogen compounds for the plant in exchange for a carbon source (photosynthate) for energy and growth of the bacteria.
Pseudomonas diazotrophicus is a gram-negative, rod-shaped, nitrogen-fixing bacterium isolated from the roots of rice (Oryza sativa) (Watanabe et al., Can. J. Microbiol., 33: 670-678 (1987). The present invention is based, at least in part, upon the discovery that Pseudomonas diazotrophicus can function as a free-living microorganism and provide biological nitrogen fixation for a large variety of plants.